The invention relates to the field of immunology and the development of adaptive immunity. More specifically the invention involves B7-related co-stimulatory molecules that are involved in the T lymphocyte response.
Induction of a T lymphocyte response is a critical initial step in a host""s immune response. Activation of T cells results in cytokine production by T cells, T cell proliferation, and generation of T-cell-mediated effector functions.
The cytokines are a diverse group of structurally dissimilar and genetically unrelated molecules. Cytokines serve as crucial intercellular-signaling molecules that are responsible for the multidirectional communication among immune and inflammatory cells engaged in host defense, repair, and restoration of homeostasis, as well as among other somatic cells in the connective tissues, skin, nervous system, and other organs. More particularly, this diverse group of intercellular-signaling proteins regulates local and systemic immune and inflammatory responses as well as wound healing, hematopoiesis, and many other biological processes.
Each cytokine is secreted by particular cell types in response to a variety of stimuli and produces a characteristic constellation of effects on the growth, motility, differentiation, or function of its target cells. In fact, cytokines regulate one another""s production and activities. Other types of biological mediators, such as corticosteroids and prostaglandins, have agonistic or antagonistic effects on cytokine activities.
Interleukin-2 (IL-2) is an autocrine and paracrine growth factor that is secreted by activated T lymphocytes. IL-2 is a critical immunoregulatory cytokine as it is essential for clonal T-cell proliferation, is involved in cytokine production, and influences the functional properties of B cells, macrophages, and NK cells. IL-2 enhances proliferation and antibody secretion by normal B cells. However, the concentration required for the B-cell response is two- to three-fold higher than is required to obtain T-cell responses. Higher concentrations of IL-2 can also activate neutrophils. IL-2 exhibits a short half-life in the circulation. Thus, it generally acts only on the cell that secreted it or on cells in the immediate vicinity.
The IL-2 receptor is not expressed in resting T cells but is induced to maximal levels within two or three days after the cells become activated. A decline in receptor expression occurs up to 6-10 days after activation. This transient nature of IL-2 receptor expression maintains the cyclical, self-limiting pattern of normal T-cell growth in vivo.
During the course of an immune response, T cells differentiate into Th phenotypes defined by their pattern of cytokine secretion and immunomodulatory properties (Abbas et al. (1996) Nature 383:787). Th cells are composed of at least two distinct subpopulations, termed Th1 and Th2 cell subpopulations (Mosmann et al. (1989) Ann. Rev. Immunol. 7:145; Del Prete et al. (1991) J. Clin. Invest. 88:346; Wiernenga et al. (1990) J. Immunol. 144:4651; Yamamura et al. (1991) Science 254:277; Robinson et al. (1993) J. Allergy Clin. Immunol. 92:313). Th1 and Th2 cells appear to function as part of the different effector functions of the immune system (Mosmann et al. (1989) Ann. Rev. Immunol. 7:145). Specifically, Th1 cells direct the development of cell-mediated immunity, triggering phagocyte-mediated host defenses, and are associated with delayed hypersensitivity. Accordingly, infections with intracellular microbes tend to induce Th1-type responses. Th2 cells drive humoral immune responses, which are associated with, for example, defenses against certain helminthic parasites, and are involved in antibody and allergic responses.
Th1 cells secrete interleukin-2 (IL-2), interferon-xcex3 (IFN-xcex3), and tumor neucrosis factor-xcex1 (TNF-xcex1). These cytokines enhance inflammatory cell-mediated responses and have a pathogenic role in the development of autoimmune disease. Th2 cells secrete interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-10 (IL-10), and interleukin-13 (IL-13). These cytokines suppress inflammatory responses while potentiating humoral immunity and control and reverse disease evolution (Scott et al. (1994) Immunity 1:73; Smith et al. (1998) J. Immunol. 160:4841; Abbas et al. (1996) Nature 383:787). The different type of cytokines released upon stimulation has been demonstrated to be central to disease evolution (Chu and Londei (1996) J. Immunol. 157:2685; Hsieh et al. (1993) Science 260:547).
T-cell activation requires two signals. The first is an antigen-specific signal, often called a primary activation signal, which results from stimulation of a T-cell receptor present on the surface of the T cell. This antigen-specific signal is usually in the form of an antigenic peptide bound either to a major histocompatibility complex (hereafter MHC) class I protein or an MHC class II protein present on the surface of an antigen presenting cell (hereafter APC). For a review see Germain (1986) Nature 322:687-691.
In addition to an antigen-specific primary activation signal, T cells also require a second, non-antigen specific signal, to induce T-cell proliferation and/or cytokine production. This phenomenon has been termed co-stimulation (Mueller et al. (1989) Annu. Rev. Immunol. 7:445-480). This xe2x80x9ctwo signalxe2x80x9d concept explains why adaptive immunity is elicited by microbes and not by self-antigens, which do not induce second signals.
Like the antigen-specific signal, the co-stimulatory signal is triggered by a molecule on the surface of the antigen presenting cell (APC). The B7 molecules are an emerging family of immunoglobulin co-stimulatory molecules, first identified on B lymphocytes (Linsley et al. (1990) Proc. Natl. Acad. Sci. 87:5031-5035). Both B7-1 (CD80) and B7-2 (CD86) bind to the T cell receptors CD28 and CTLA4, resulting in co-stimulation of the T cell (Peach et al. (1995) J. Biol. Chem. 270:21181-21187; Fargeas et al. (1995) J. Exp. Med. 182:667-675; Bajorath et al. (1994) Protein Sci. 3:2148-2150; U.S. Pat. No. 5,942,607; and PCT Application No. WO 96/40915). Depending upon which receptor is bound, the activated T-cell immune response is enhanced (CD28) or inhibited (CTLA4) in a negative feedback loop. Additional B7 homologs have been identified including B7-H1, and B7RP-1 and its mouse ortholog B7h (Swallow et al. (1999) Immunity 11:423-432; Dong et al. (1999) Nature Med. 5:1365-1369; Yoshinaga et al. (1999) Nature 402:827-832). Although both B7RP-1 and B7-H1 co-stimulate T-cell proliferation, neither of these molecules binds to either CD28 or CTLA4 (Abbas and Sharpe (1999) Nature Med. 5:1345-1346; Yoshinaga et al. (1999) Nature 402:827-832). Unlike B7-1 and B7-2, B7-H1 has little effect on IL-2 production, but considerably increases T-cell production of IL-10, a B-cell differentiation factor that inhibits macrophages and cell-mediated immunity.
Ligation of the CD28 family member ICOS (inducible co-stimulator) increases IL-10 production. B7RP-1 has been shown to bind to this receptor (Yoshinaga et al. (1999) Nature 402:827-832) while B7-H1 does not appear to bind to ICOS (Dong et al. (1999) Nature Med. 5:1365-1369), although this result is not definitive. Like CD28, ICOS enhances all basic T-cell responses to a foreign antigen, namely, proliferation, secretion of lymphokines, up-regulation of molecules that mediate cellxe2x80x94cell interaction, and effective help for antibody secretion by B-cell. Unlike the constitutively expressed CD28, ICOS has to be de novo induced on the T-cell surface, does not up-regulate the production of IL-2, but superinduces the synthesis of IL-10 (Hutloff et. al. (1999) Nature 397:263-266). The inducible expression of ICOS shortly after T-cell activation indicates that ICOS may be particularly important in providing co-stimulatory signals to activated T cells, in contrast to CD28, which is essential in the activation and differentiation of naxc3xafve T cells (McAdam et al. (1998) Immunol. Rev. 165:231-247). ICOS may down-regulate immune responses by stimulating development of regulatory T cells, which normally function to control the injurious side effects of cell-mediated immunity. As ICOS signaling induces IL-10, which can also down-regulate B7-1 and B7-2 expression (Ding et al. (1993) J. Immunol. 151:1224-1234), ICOS co-stimulation may indirectly reduce or inhibit B7 expression and thereby inhibit B7-mediated CD28 co-stimulation. Therefore, whereas B7-1 and B7-2 function in the initiation and development of immune responses, B7RP-1 and B7-H1 may function to return the immune system to its resting state.
Another receptor belonging to the immunoglobulin gene superfamily, designated PD-1, also appears to be involved in the negative regulation of certain immune responses. PD-1 knockout mice develop Lupus-like autoimmune diseases (Nishimura et al. (1999) Immunity 11:141-151). In addition, the identification of a novel member of the B7 family (PD-L) that binds to the PD-1 receptor but not CD28, CTLA4, or ICOS has been reported (Freeman et al. (2000) FASEB J. 14(6):Abstract 153.34).
The profile of the natural immune response, specifically cytokine production, may determine the phenotype of the subsequent immune response. Therefore, methods are needed to regulate an immune response. There is great interest in the possibility that in disease situations in which antigens are either unknown or difficult to manipulate, immune responses may be either enhanced or terminated by manipulating the co-stimulation signals such as those signals affected by the B7 family of proteins. For example, modulating the co-stimulation signals may promote tumor immunity and reduce graft rejection, autoimmune, inflammatory, and infectious diseases (Abbas and Sharpe (1999) Nature Med. 5:1345-1346; Schweiter and Sharpe (1998) J. Immunol. 161:2762-2771; Wallace et al. (1994) Transplantation 58:602; Sayegh (1995) J. Exp. Med. 181:1869; Lenschow et al. (1995) J. Exp. Med. 181:1145; Finck et al. (1994) Science 265:1225; Cross et al. (1995) J. Clin. Invest. 95:2783; Perrin et al. (1995) J. Immunol. 154:1481; Corry et al. (1994) J. Immunol. 153:4142; U.S. Pat. Nos. 5,968,510, 5,861,310, and 5,521,288; and PCT Application No. WO 90/05541 and European Patent No. EP445228B 1).
Isolated nucleic acid molecules, hB7-H2 long (hB7-H21) and hB7-H2 short (hB7-H2s), corresponding to B7-like nucleic acid sequences are provided. Additionally, amino acid sequences corresponding to the polynucleotides are encompassed. In particular, the present invention provides for isolated nucleic acid molecules comprising nucleotide sequences encoding the amino acid sequences shown in SEQ ID NO:2 and SEQ ID NO:4, the nucleotide sequence encoding the DNA sequence deposited in a bacterial host as ATCC Accession Number PTA-2084, or the nucleotide sequence encoding the DNA sequence deposited in a bacterial host as ATCC Accession Number PTA-2085. Further provided are B7-like polypeptides having amino acid sequences encoded by the nucleic acid molecules described herein.
The present invention also provides vectors and host cells for recombinant expression of the nucleic acid molecules described herein, as well as methods of making such vectors and host cells and for using them for production of the polypeptides or peptides of the invention by recombinant techniques.
Another aspect of this invention features isolated or recombinant B7-like proteins and polypeptides. Preferred B7-like proteins and polypeptides possess at least one biological activity possessed by naturally occurring B7-like proteins.
Variant nucleic acid molecules and polypeptides substantially homologous to the nucleotide and amino acid sequences set forth in the Sequence Listing are encompassed by the present invention. Additionally, fragments and substantially homologous fragments of the nucleotide and amino acid sequences are provided.
Antibodies and antibody fragments that selectively bind B7-like polypeptides and fragments are provided. Such antibodies are useful in detecting B7-like polypeptides as well as in regulating the T-cell immune response and cellular activity.
The B7-like molecules of the present invention are useful for modulating immune responses. The molecules of the invention are useful for the treatment and diagnosis of T-lymphocyte-related disorders, including, but not limited to, atopic conditions, such as asthma and allergy, including allergic rhinitis, psoriasis, the effects of pathogen infection, chronic inflammatory diseases, autoimmune diseases, graft rejection, graft versus host disease and neoplasia. Compositions of the invention are useful in the treatment and diagnosis of disorders related to bone-metabolism, and in the treatment and diagnosis of cancers such as B7 lymphomas, carcinomas, and T cell leukemias, and useful for treatment of viral diseases and cancers such as herpes, Kaposi""s sarcoma, genital warts, hairy cell leukemia, melanoma, and renal cell carcinoma.
In addition, the molecules of the invention are useful as modulating agents in a variety of cellular processes including growth promoting activity, particularly the antigen-independent proliferation of T helper cell clones, and direct effects on normal hemopoietic progenitors, human T cells, B cells, thymocytes, thymic lymphomas, and neuronal cell lines.
This invention provides isolated nucleic acid molecules encoding B7-like proteins or biologically active portions thereof, as well as nucleic acid fragments suitable as primers or hybridization probes for the detection of B7-like-encoding nucleic acids.
In another aspect, the present invention provides a method for detecting the presence of B7-like activity or expression in a biological sample by contacting the biological sample with an agent capable of detecting an indicator of B7-like activity such that the presence of B7-like activity is detected in the biological sample.
In yet another aspect, the invention provides a method for modulating B7-like activity comprising contacting a cell with an agent that modulates (inhibits or stimulates) B7-like activity or expression such that B7-like activity or expression in the cell is modulated. In one embodiment, the agent is an antibody that specifically binds to a B7-like protein. In another embodiment, the agent modulates expression of B7-like proteins by modulating transcription of a B7-like gene, splicing of a B7-like mRNA, or translation of a B7-like mRNA. In yet another embodiment, the agent is a nucleic acid molecule having a nucleotide sequence that is antisense to the coding strand, or to a portion thereof, of the B7-like mRNA or the B7-like gene.
In another aspect, the invention provides a method for identifying a compound that binds to or modulates the activity of a B7-like protein and/or its binding partner. In general, such methods entail measuring a biological activity of a B7-like protein in the presence and absence of a test compound and identifying those compounds that alter the activity of the B7-like protein.
In one embodiment, the methods of the present invention are used to treat a subject having a disorder that involves B7-like protein activity or nucleic acid expression by administering an agent that is a B7-like modulator to the subject. In one embodiment, the B7-like modulator is a B7-like protein. In another embodiment, the B7-like modulator is a B7-like nucleic acid molecule. In other embodiments, the B7-like modulator is a peptide, peptidomimetic, or other small molecule. In another embodiment the B7-like modulator is an antibody specific for B7-like proteins.
The present invention also provides a diagnostic assay for identifying the presence or absence of a genetic lesion or mutation characterized by at least one of the following: (1) aberrant modification or mutation of a gene encoding a B7-like protein; (2) misregulation of a gene encoding a B7-like protein; and (3) aberrant post-translational modification of a B7-like protein, wherein a wild-type form of the gene encodes a protein with a B7-like activity.
The invention also features methods for identifying a compound that modulates the expression of B7-like genes by measuring the expression of the B7-like sequences in the presence and absence of the compound.